Quantitative assessment of thermal denaturation of bovine alpha-lactalbumin via low-intensity ultrasound, HPLC, and DSC

The degree of irreversible aggregation and the aggregation velocity constant of alpha-lactalbumin (alpha-la) were determined by three methods based on different principles: low-intensity ultrasound as a novel method for this purpose, DSC, and HPLC. The denaturation process of alpha-la causes a decrease in the ultrasonic velocity due to the conformation change of alpha-la molecules. This decrease is a function of the concentration of native alpha-la in the sample. A linear correlation was found between the degree of aggregation of alpha-la determined by these three methods. There is no significant difference between the aggregation velocity constants determined by the three methods. The results show that the ultrasonic method is capable of quantifying the degree of aggregation of a protein, offering an alternative method.